Acidification is required for calcium and magnesium concentration measurements in equine urine

Acidification of equine urine to promote dissociation of ion complexes is a common practice for urine ion concentration measurements. The objective of this study was to evaluate the effect of acidification and storage after acidification on calcium (Ca), magnesium (Mg) and phosphate (P) concentrations and on fractional excretion (FE) of these electrolytes. Thirty-two fresh equine urine samples were analysed between December 2016 and July 2020. Complete urinalysis (stick and sediment) was performed on all samples. Ca, Mg, P and creatinine concentrations were measured in supernatant of centrifuged native urine, urine directly centrifuged after acidification and urine centrifuged 1 hour after acidification. Urine was acidified with hydrochloric acid to reach a pH of 1-2. Ca, Mg, P and creatinine concentrations were also measured in blood plasma, and fractional excretion of each electrolyte was calculated. Equality of medians was tested with Friedman tests and Bland-Altman bias plots were used to show the agreement between conditions.

Acidification had a statistically significant effect on Ca and Mg concentrations, FE_Ca and FE_Mg. Bland-Altman plot revealed a strong positive proportional bias between Ca concentration in native and acidified urine with a mean bias of 17.6 mmol/l. For Mg concentration, the difference between native and acidified urine was small with a mean bias of 1.8 mmol/l. The increase in FE_Ca was clinically relevant. Storage of acidified urine had no effect on any of the measured ion concentrations. All P concentrations in native urine samples were below the detection limit of the assay and statistical analysis and calculation of FE_P was not possible.

Urine acidification is essential for accurate measurement of Ca and Mg concentrations and therefore FE calculations in equine urine. Storage time of 1 hour after acidification does not significantly change Ca and Mg concentrations.